Biological research and applications in biotechnology often require cell lines that express high levels of a given gene product. As an increasing number of genes are isolated and developed for the production of a wide array of useful polypeptide drugs, there is an increasing need to enhance the efficiencies and economies of manufacture. Much of the effort in this direction has been directed to the use of strong promoters, enhancers, high copy number plasmids, and amplification using an amplifiable gene.
While amplification appears to be an attractive approach, nevertheless it has many limitations. The amplification process has normally involved ligating in tandem the amplifiable gene and the gene of interest, where each of the genes has an independent transcriptional initiation region. For the most part, this approach while showing some promise has not proven to be as useful a might have been hoped. The level of amplification has been limited. In addition, the tandem sequences have been unstable in that in the absence of continuous selective pressure, copies of the gene may be looped out and lost. Since selective pressure normally reduces the viability of the cells, there is an interest in being able to develop systems to provide for stable enhancement of expression of a desired gene.